title: MicroRNA-29a ameliorates glucocorticoid-induced suppression of osteoblast differentiation by regulating β-catenin acetylation.
abstract: Excess glucocorticoid treatment induces loss of osteoblast differentiation. Post-translational modification of β-catenin reportedly regulates osteogenic activities in bone cells. This study was undertaken to test whether miR-29a signaling regulates the acetylation status of β-catenin in the glucocorticoid-mediated osteoblast dysfunction. Murine osteoblast cultures were incubated under osteogenic conditions with or without supraphysiological glucocorticoid, miR-29a precursor, antisense oligonucleotides or histone deacetylase 4 (HDAC4) RNA interferences. Osteoblast differentiation was determined by alkaline phosphatase activity, calcium deposition, and von Kossa stain. β-Catenin acetylation and miR-29a transcription were detected by immunoblotting, chromatin immunoprecipitation and quantitative PCR. Protein interaction was detected by fluorescence protein ligation assay. Supraphysiological glucocorticoid treatment repressed osteoblast differentiation and induced loss of miR-29a expression and acetylated β-catenin levels in osteoblast cultures. Gain of miR-29a function attenuated the deleterious effects of glucocorticoid on osteogenic gene expression and mineralized nodule formation, whereas knockdown of miR-29a signaling accelerated loss of osteoblast differentiation capacity. miR-29a reduced HDAC4 signaling and attenuated the glucocorticoid-mediated β-catenin deacetylation and ubiquitination and restored nuclear β-catenin levels. Glucocorticoid-induced loss of miR-29a signaling occurred through transcriptional and translational regulation. Interruption of HDAC4 signaling attenuated the glucocorticoid-induced hypoacetylation of histone H3 at lysine 9 (H3K9Ac) and restored the enrichment of H3K9Ac in miR-29a proximal promoter region and miR-29a transcription in cell cultures. Taken together, excess glucocorticoid-induced loss of miR-29a signaling accelerates β-catenin deacetylation and ubiquitination that impairs osteogenic activities of osteoblast cultures. miR-29a and HDAC4 reciprocal regulation of H3K9 acetylation contributes to the acetylation status of β-catenin and miR-29a expression. Enhancement of miR-29a signaling is an alternative strategy for protecting against the adverse actions of excess glucocorticoid on differentiation capacity of osteogenic cells.
authors.affiliation: Department of Orthopedic Surgery, Kaohsiung Chang Gung Memorial Hospital, Taiwan; Department of Medical Research, Kaohsiung Chang Gung Memorial Hospital, Taiwan.
authors.forename: Jih-Yang
authors.initials: JY
authors.lastname: Ko
authors.forename: Pei-Chin
authors.initials: PC
authors.lastname: Chuang
authors.forename: Ming-Wen
authors.initials: MW
authors.lastname: Chen
authors.forename: Huei-Ching
authors.initials: HC
authors.lastname: Ke
authors.forename: Shin-Long
authors.initials: SL
authors.lastname: Wu
authors.forename: Yu-Hsuan
authors.initials: YH
authors.lastname: Chang
authors.forename: Yu-Shan
authors.initials: YS
authors.lastname: Chen
authors.forename: Feng-Sheng
authors.initials: FS
authors.lastname: Wang
mesh_terms: D000107:Acetylation / Q000187:drug effects; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects* / Q000235:genetics; D002460:Cell Line; D005938:Glucocorticoids / Q000494:pharmacology*; D006655:Histone Deacetylases / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D051379:Mice; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D034622:RNA Interference / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D012886:Skull / Q000166:cytology; D014158:Transcription, Genetic / Q000187:drug effects; D054875:Ubiquitination / Q000187:drug effects; D051176:beta Catenin / Q000378:metabolism*
chemical_list: C501127:Dkk1 protein, mouse; D005938:Glucocorticoids; D036341:Intercellular Signaling Peptides and Proteins; C547014:MIRN29 microRNA, mouse; D035683:MicroRNAs; D016376:Oligonucleotides, Antisense; D051176:beta Catenin; C498618:Hdac5 protein, mouse; D006655:Histone Deacetylases
keywords: Acetylation; Glucocorticoid; Osteoblasts; miR-29a; β-Catenin